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The DNA-Encoded Nucleosome Organization of a Eukaryotic Genome

ABSTRACT: Nucleosome organization is critical for gene regulation. In living cells, this organization is determined by multiple factors, including the action of chromatin remodelers, competition with site-specific DNA-binding proteins, and the DNA sequence preferences of the nucleosomes themselves. However, it has been difficult to estimate the relative importance of each of these mechanisms in vivo, because in vivo nucleosome maps reflect the combined action of all influencing factors. Here, we determine the importance of DNA sequence preferences experimentally by measuring the genome-wide occupancy of nucleosomes assembled on purified yeast genomic DNA. The resulting map, in which nucleosome occupancy is governed only by the intrinsic sequence preferences of nucleosomes, is remarkably similar to in vivo nucleosome maps generated in three different growth conditions. In vitro, nucleosome depletion is evident at many transcription factor binding sites and around gene start and end sites, suggesting that nucleosome depletion at these sites in vivo is partially encoded in the genome. We confirm these results with a micrococcal nuclease-independent experiment that measures the relative affinity of nucleosomes for ~40,000 double-stranded 150bp oligonucleotides. Using our in vitro data, we devise a computational model of nucleosome sequence preferences that is significantly correlated with in vivo nucleosome occupancy in C. elegans. Our results indicate that the intrinsic DNA sequence preferences of nucleosomes play a central role in determining the organization of nucleosomes in vivo.

OTHER RELATED OMICS DATASETS IN: PRJNA109407PRJNA110649

ORGANISM(S): synthetic construct Mus musculus Saccharomyces cerevisiae Homo sapiens

PROVIDER: GSE13622 | GEO | 2008/12/18

SECONDARY ACCESSION(S): PRJNA110649

REPOSITORIES: GEO

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Project description:Human CMV (hCMV) establishes lifelong infections in most of us, causing developmental defects in human embryos and life-threatening disease in immunocompromised individuals. During productive infection, the viral >230,000-bp dsDNA genome is expressed widely and in a temporal cascade. The hCMV genome does not carry histones when encapsidated but has been proposed to form nucleosomes after release into the host cell nucleus. Here, we present hCMV genome-wide nucleosome occupancy and nascent transcript maps during infection of permissive human primary cells. We show that nucleosomes occupy nuclear viral DNA in a nonrandom and highly predictable fashion. At early times of infection, nucleosomes associate with the hCMV genome largely according to their intrinsic DNA sequence preferences, indicating that initial nucleosome formation is genetically encoded in the virus. However, as infection proceeds to the late phase, nucleosomes redistribute extensively to establish patterns mostly determined by nongenetic factors. We propose that these factors include key regulators of viral gene expression encoded at the hCMV major immediate-early (IE) locus. Indeed, mutant virus genomes defcient for IE1 expression exhibit globally increased nucleosome loads and reduced nucleosome dynamics compared with WT genomes. The temporal nucleosome occupancy differences between IE1-defcient and WT viruses correlate inversely with changes in the pattern of viral nascent and total transcript accumulation. These results provide a framework of spatial and temporal nucleosome organization across the genome of a major human pathogen and suggest that an hCMV major IE protein governs overall viral chromatin structure and function. [i] H3 ChIP-chip measurements of WT human cytomegalovirus (strain TB40E) following infection of MRC-5 cells. WT 8 hours postinfection, with corresponding mock IgG input control: 3 replicates; WT virus, 48 hours postinfection with corresponding mock IgG input contro: 3 replicates. [ii] MNase-chip measurements of WT and dlIE1 human cytomegalovirus nucleosomes following infection of MRC-5 cells. WT 8 hours postinfection, with corresponding sonicated DNA input control: 6 replicates; WT virus 48 hours postinfection, with corresponding sonicated DNA input control: 6 replicates; WT 96 hours postinfection, with corresponding sonicated DNA input control: 2 replicates; dlIE1 virus 8 hours postinfection, with corresponding sonicated DNA input control: 2 replicates; dlIE1 virus 96 hours postinfection, with corresponding sonicated DNA input control: 2 replicates. [iii] Total and nascent RNA measurements of WT and dlIE1 human cytomegalovirus transcripts following infection of MRC-5 cells. WT 8 hours postinfection: 2 replicates; WT virus 96 hours postinfection: 2 replicates; dlIE1 8 hours postinfection: 2 replicates; dlIE1 virus 96 hours postinfection: 2 replicates; sonicated DNA input control: 10 replicates.

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The DNA-Encoded Nucleosome Organization of a Eukaryotic Genome

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